Tuning Riboswitch‐Mediated Gene Regulation by Rational Control of Aptamer Ligand Binding Properties

Riboswitch‐mediated control of gene expression depends on ligand binding properties (kinetics and affinity) of its aptamer domain. A detailed analysis of interior regions of the aptamer, which affect the ligand binding properties, is important for both understanding natural riboswitch functions and for enabling rational design of tuneable artificial riboswitches. Kinetic analyses of binding reaction between flavin mononucleotide (FMN) and several natural and mutant aptamer domains of FMN‐specific riboswitches were performed. The strong dependence of the dissociation rate (52.6‐fold) and affinity (100‐fold) on the identities of base pairs in the aptamer stem suggested that the stem region, which is conserved in length but variable in base‐pair composition and context, is the tuning region of the FMN‐specific aptamer. Synthetic riboswitches were constructed based on the same aptamer domain by rationally modifying the tuning regions. The observed 9.31‐fold difference in the half‐maximal effective concentration (EC₅₀) corresponded to a 11.6‐fold difference in the dissociation constant (K_D) of the aptamer domains and suggested that the gene expression can be controlled by rationally adjusting the tuning regions.     

Toehold‐Mediated Displacement of an Adenosine‐Binding Aptamer from a DNA Duplex by its Ligand

DNA is increasingly used to engineer dynamic nanoscale circuits, structures, and motors, many of which rely on DNA strand‐displacement reactions. The use of functional DNA sequences (e.g., aptamers, which bind to a wide range of ligands) in these reactions would potentially confer responsiveness on such devices, and integrate DNA computation with highly varied molecular stimuli. By using high‐throughput single‐molecule FRET methods, we compared the kinetics of a putative aptamer–ligand and aptamer–complement strand‐displacement reaction. We found that the ligands actively disrupted the DNA duplex in the presence of a DNA toehold in a similar manner to complementary DNA, with kinetic details specific to the aptamer structure, thus suggesting that the DNA strand‐displacement concept can be extended to functional DNA–ligand systems.     

Protein-protein Interaction Between Domains of PDZ and BAR from PICK1

IntroductionProtein kinase Cα(PKCα) widely existed in thecells of human being and animals is one kind of serine/threonine kinase that can be rapidly activated by a vari-ety of factors such as Ca2 , cAMP and hormone, re-sulting in the signal transmissio…     

氮、硫双齿配体钯促进的Suzuki-Miyaura交叉偶联反应

钯配合物促进的Suzuki-Miyaura交叉偶联反应为构建碳-碳骨架的重要方法,主要用于药物化学、天然产物、材料学及超分子配体等领域。以2-噻吩甲醛、脂肪族/芳香族伯胺、取代肼及羟胺等为原料,合成了9种2-噻吩甲醛亚胺类N,S-双齿配体（3a~3i）,其结构经¹H NMR, ¹³C NMR和HR-MS（APCI）表征。并以N-环己基-2-噻吩甲亚胺(3b)为例,研究了配体对钯促进的Suzuki-Miyaura交叉偶联反应的催化性能。结果表明：在反应体系Pd(OAc)₂/3b/Cs₂CO₃/DMF中,苯硼酸与芳溴的交叉偶联反应的分离产率达到95%。     

Characterization of Ligand Binding by Saturation Transfer Difference NMR Spectroscopy

Fast identification of binding activity directly from mixtures of potential ligands is possible with the NMR method described, which is based on saturation transfer to molecules in direct contact to a protein. In addition, the ligand's binding epitope is easily identified. High sensitivity and ease of use are the principal advantages of this method. The picture shows the normal 1D NMR spectrum of a mixture and the spectrum obtained by applying the STD method, which exclusively shows signals from molecules with binding affinity.     

Iron Pentacarbonyl Promoted Addition of CO and MeOH to 1,4-Disubstituted-1,3-butadiyne and Formation of Vinylallyl and Butatriene Ligand Systems

Abstract  Photochemical reaction of methanol solution containing 1,4-diferrocenyl- or 1,4-diphenyl-1,3-butadiynes and iron pentacarbonyl into which CO was constantly bubbled, yielded diiron hexacarbonyl complexes of cumulene ligand systems, [η¹: η³-{RCHC₂CR(COOMe)}Fe₂(CO)₆] (1; E, R = Fc, 2; Z, R = Fc, 5; E, R = Ph, 6; Z, R = Ph) and [η³: η³-{RCHC₂CR(COOMe)}Fe₂(CO)₆] (3; E, R = Fc, 7; E, R = Ph), formed by 1,4-addition of –COOMe and –H to the butadiynes. Additionally, diferrole, [Fe(CO)₄{C(O)CC(Fc)C(O)}₂],4 was obtained in minor quantity. Compounds 1, 2, 5 and 6 contain vinylallyl carbon framework which is stabilized by MeOC=O → Fe bond along with η¹: η³ coordinated Fe₂(CO)₆ unit. Compounds 3 and 7 contain butatriene units which are stabilized by η³: η³ coordinated Fe₂(CO)₆ unit. Characterization of the new compounds was carried out by IR and ¹H and ¹³C NMR spectroscopy and by mass spectrometry. Molecular structures of 2–7 were established by single crystal X-ray diffraction methods. Graphical Abstract  Diiron hexacarbonyl complexes of cumulene ligand systems, [η¹: η³ {RCHC₂CR(COOMe)}] (1; E, R = Fc, 2; Z, R = Fc, 5; E, R = Ph, 6; Z, R = Ph) and [η³: η³-{RCHC₂CR(COOMe)}] (3; E, R = Fc, 7; E, R = Ph) were obtained from photochemical reactions between Fe(CO)₅, CO and methanol. Yield of the minor product, the diferrole, 4, was improved when the photoreaction was carried out in hexane in place of methanol      

Catalytic Dinitrogen Reduction at the Molybdenum Center Promoted by a Bulky Tetradentate Phosphine Ligand

Stoichiometric reduction of N₂ at a Mo center stabilized by a bulky tetradentate phosphine ligand (${{\rm PP}{{{\rm Cy}\hfill \atop 3\hfill}}}$ ) allowed isolation of Mo–imidoamine and Mo–imido complexes. Both complexes as well as the Mo^II precursor are equally suitable catalysts for the synthesis of NTMS₃ (TMS=trimethylsilyl) from N₂, TMSCl, and electron sources. Mechanistic studies prove the involvement of a TMS radical at least in one of the catalytic steps.     

Carbon-Carbon Bond Formation Reaction Promoted by Cadmium In Aqueous Media

Promoted by metallic cadmium allylilic and propargyl bromides react smoothly with aldehydes in aqueous DMF to give homoallylic and homopropargyl alcohols in moderate to good yields. It can also promote pinacol coupling of aromatic aldehydes. The metallic cadmium is produced in situ by the reduction of CdCl₂ H₂O with samarium metal.     

An Aqueous Electrolyte Gated Artificial Synapse with Synaptic Plasticity Selectively Mediated by Biomolecules

The emulation of functions and behaviors of biological synapses using electronic devices has inspired the development of artificial neural networks (ANNs) in biomedical interfaces. Despite the achievements, artificial synapses that can be selectively responsive to non-electroactive biomolecules and directly operate in biological environments are still lacking. Herein, we report an artificial synapse based on organic electrochemical transistors and investigate the selective modulation of its synaptic plasticity by glucose. The enzymatic reaction between glucose and glucose oxidase results in long-term modulation of the channel conductance, mimicking selective binding of biomolecules to their receptors and consequent long-term modulation of the synaptic weight. Moreover, the device shows enhanced synaptic behaviors in the blood serum at a higher glucose concentration, which suggests its potential application in vivo as artificial neurons. This work provides a step towards the fabrication of ANNs with synaptic plasticity selectively mediated by biomolecules for neuro-prosthetics and human-machine interfaces.     

Ligand Binding and Release Investigated by Contact-Guided Iterative Multiple Independent Molecular Dynamics Simulations

Binding and releasing ligands are critical for the biological functions of many proteins, so it is important to determine these highly dynamic processes. Although there are experimental techniques to determine the structure of a protein-ligand complex, it only provides a static picture of the system. With the rapid increase of computing power and improved algorithms, molecular dynamics (MD) simulations have diverse of superiority in probing the binding and release process. However, it remains a great challenge to overcome the time and length scales when the system becomes large. This work presents an enhanced sampling tool for ligand binding and release, which is based on iterative multiple independent MD simulations guided by contacts formed between the ligand and the protein. From the simulation results on adenylate kinase, we observe the process of ligand binding and release while the conventional MD simulations at the same time scale cannot.     

Binding of Dopamine to Alpha-Synuclein is Mediated by Specific Conformational States

Parkinson’s disease is the second most common neurodegenerative disorder, in which both alpha-synuclein (α-syn) and dopamine (DA) have a critical role. α-Syn is known to be natively unstructured in equilibrium with subpopulations of more compact structures. It is these compact structures that are thought to be linked to amyloid formation. In the presence of DA, α-syn yields a diverse range of SDS-resistant, non-amyloid oligomers, however the precursor state conformation has not been established. Here, three DA molecules have been observed to bind per α-syn monomer by electrospray-ionization-ion mobility spectrometry-mass spectrometry (ESI-IMS-MS). Each of these DA molecules binds exclusively to the extended conformation of α-syn, and binding is not observed in the compact state of the protein. Measurements of collisional cross sectional areas show that the incremental uptake of DA pushes the protein towards a highly extended population, becoming fully populated upon the binding of three DA ligands. Tyrosine (Tyr) as a closely related structural analog, exhibited limited binding to the protein as compared with DA, with a maximum of two ligands being observed. Those Tyr ligands that do bind were observed as adducts to the extended conformation akin to DA. These findings suggest DA is able to modulate α-syn self-assembly by inducing the population of a highly extended state.

DNA小沟结合二脒:水分子的识别作用

采用分子动力学模拟了DB921-DNA复合物, 通过7 ns的模拟研究表明: DB921一端的氨基氮原子与一个水分子形成氢键, 同时, 水分子又与DNA的5位A碱基的氮原子形成一个氢键. 水分子在DB921与DNA小沟结合中起了桥连的作用, 使得直线型的芳香二脒化合物DB921通过水桥与DNA小沟结合, 水分子诱导DB921分子与DNA的小沟域构型相适应, 与DNA小沟域的AATTC碱基有较强的结合作用. 在分子水平上提供了DB921与双螺旋DNA相互作用的结构及复合物的动态变化情况, 指出水分子在DNA小沟结合二脒化合物中的识别作用, 为设计出更高生物活性的DNA小沟结合剂提供一定的理论依据.     

Inhibition of Ligand Binding Ability of Three Porphyrins by an Organic Effector

A stimulus‐responsive receptor 1 was designed and prepared to control the ligand‐binding ability of three active sites, two zinc tetraphenylporphyrin units (P1) and one zinc diethynyldiphenylporphyrin unit (P2), with one effector molecule 2 . Bulky hexarylbenzene units were incorporated as shielding panels in the middle of the flexible side arms of 1 . Spectroscopic titrations indicated that a stable supramolecular complex 1 ? 2 (K _{1 ? 2} =6.7×10⁶ m ^?1) was produced by the cooperative formation of multiple hydrogen and coordination bonds. As a result, the binding of a ligand to P1 was inhibited by 2 in a competitive manner. Additionally, the formation of 1 ? 2 brought about conformational restriction of the side arms to cover both faces of P2 with the shielding panels. The binding constant of 4‐phenylpyridine with P2 in 1 ? 2 decreased to 8.9 % of that in 1 . Namely, the ligand‐binding ability of P2 was inhibited according to an allosteric mechanism.     

Domain Analysis and Motif Matcher (DAMM): A Program to Predict Selectivity Determinants in Monosiga brevicollis PDZ Domains Using Human PDZ Data

Choanoflagellates are single-celled eukaryotes with complex signaling pathways. They are considered the closest non-metazoan ancestors to mammals and other metazoans and form multicellular-like states called rosettes. The choanoflagellate Monosiga brevicollis contains over 150 PDZ domains, an important peptide-binding domain in all three domains of life (Archaea, Bacteria, and Eukarya). Therefore, an understanding of PDZ domain signaling pathways in choanoflagellates may provide insight into the origins of multicellularity. PDZ domains recognize the C-terminus of target proteins and regulate signaling and trafficking pathways, as well as cellular adhesion. Here, we developed a computational software suite, Domain Analysis and Motif Matcher (DAMM), that analyzes peptide-binding cleft sequence identity as compared with human PDZ domains and that can be used in combination with literature searches of known human PDZ-interacting sequences to predict target specificity in choanoflagellate PDZ domains. We used this program, protein biochemistry, fluorescence polarization, and structural analyses to characterize the specificity of A9UPE9_MONBE, a M. brevicollis PDZ domain-containing protein with no homology to any metazoan protein, finding that its PDZ domain is most similar to those of the DLG family. We then identified two endogenous sequences that bind A9UPE9 PDZ with <100 μM affinity, a value commonly considered the threshold for cellular PDZ–peptide interactions. Taken together, this approach can be used to predict cellular targets of previously uncharacterized PDZ domains in choanoflagellates and other organisms. Our data contribute to investigations into choanoflagellate signaling and how it informs metazoan evolution.     

Carbon–Carbon Bond Forming Reactions Promoted by Aluminyl and Alumoxane Anions: Introducing the Ethenetetraolate Ligand

[K{Al(NON^Dipp)}]₂ (NON^Dipp=[O(SiMe₂NDipp)₂]^2?, Dipp=2,6‐iPr₂C₆H₃) reacts with CS₂ to afford the trithiocarbonate species [K(OEt₂)][Al(NON^Dipp)(CS₃)] 1 or the ethenetetrathiolate complex, [K{Al(NON^Dipp)(S₂C)}]₂ [ 3 ]₂. The dimeric alumoxane [K{Al(NON^Dipp)(O)}]₂ reacts with carbon monoxide to afford the oxygen analogue of 3 , [K{Al(NON^Dipp)(O₂C)}]₂ [ 4 ]₂ containing the hitherto unknown ethenetetraolate ligand, [C₂O₄]^4?.     

水溶液中磷酰基促进的成肽反应研究

水溶液中磷酰基促进的成肽反应研究沙耀武，巨勇，赵玉芬（清华大学化学系，北京，１０００８４）关键词磷酰化组氨酸，成肽，水溶液一些氨基酸在前生化时期的条件下，就可以由甲烷、氨和水通过高压放电生成［１，２］。如果向该反应体系中加入三氢化磷，生成的氨基酸的种...